Many procedures that are performed in biochemical laboratories involve analyses of multiple samples or materials distributed over a two-dimensional area. Examples of such procedures are screening studies performed on substances that are placed in individual wells of a multi-well plate such as a standard 96-well microtiter plate or larger plates, or on molecular species that are applied as droplets or regularly spaced spots, either microscopic in size or larger, on a solid surface. Further examples are slab-shaped electrophoresis gels in which either two-dimensional electrophoretic separations or one-dimensional separations of multiple samples in parallel have been performed. Still further examples are blotting membranes to which electrophoretically separated species in the form of spots or bands have been transferred from a slab gel. Other examples will readily occur to the skilled biochemist.
In all of these examples, detection and analysis of individual sites in the two-dimensional array can make use of radiation associated with each site. Detection can simply determine the presence or absence of particular species, or can also include quantitative determinations, either on an absolute basis or as comparisons among different sites. In some cases, the detected radiation results from the decay of radionuclides. In other cases, the detected radiation is light, which can be transmitted, absorbed, or reflected by each site, or generated by the materials at the sites themselves. Species in electrophoresis gels or blotting membranes, for example, are commonly detected by fluorescence, chemiluminescence, or bioluminescence, either as inherent characteristics of the species at the sites or as a result of treatment of the species once they are separated throughout the two-dimensional array. The treatment may include binding reactions in which energy-emitting labels are attached to the species, or irradiation of the species or the labels with excitation energy that will cause them to emit light energy, most often at different wavelengths.
In contact imaging, a two-dimensional array as discussed above is placed in close proximity to a sensor, and radiation originating from species in the array is imaged using the sensor. Examples of suitable sensors include conventional photographic film, amorphous silicon (a-Si) sensors, as well as charge-coupled devices (CCDs) and complementary metal-oxide-semiconductor (CMOS) devices. A sensor can be constructed to be planar or two-dimensional in shape, and have dimensions similar to those of the arrays it is used to image. Due to the short distance and large area of exposure between the array and sensor, contact imaging allows weak radiation sources to be detected at high spatial resolution.
Many applications of contact imaging have been optimized for radiation sources that do not require energy to be input at the time of detection. For example, conventional cassettes that hold sample arrays against photographic film are often impervious to outside light, and require that arrays contain endogenous (e.g., chemiluminescent) light sources. Such cassettes are not suitable for imaging fluorescent samples, which generally emit long-wavelength light in response to excitation with short-wavelength light. Because most fluorophores have excited-state lifetimes on the order of nanoseconds, it is impractical to excite fluorophores in an array and then enclose the array in a dark container for contact imaging.
Adapting contact imaging devices and procedures for fluorescent species presents technical challenges. The sample array must be exposed to excitation light at the time of imaging, but an image of species in the array must be formed mainly from fluorescently emitted light rather than excitation light. Contact between the array and sensor limits options for preventing excitation light from reaching the sensor.